The overall goals of the proposed research are: i) to learn more about mast cell differentiation and the generation of mast cell heterogeneity in vitro and in vivo through studies of the membrane glycosphingolipids (GSLs) of these cells; and ii) to gain fundamental knowledge about the subcellular distribution of GSLs in mast cells and about the redistribution of these molecules during mast cell activation. Mouse serosal mast cells express Forssman antigen, a neutral GSL, while mouse bone marrow culture-derive mast cells do not. The first specific aim will be to determine whether additional GSLs are selectively expressed by the populations by completing the chemical identification of their respective GSLs. These molecules will then be used as biochemical markers of the populations in the second specific aim: to determine whether or not the GSL phenotype of bone marrow culture-derive mast cells changes towards that of serosal mast cells in the presence of growth-inhibiting and maturation- inducing agents. Using specific antibodies for some of the GSLs, along with anti-mast cell monoclonal antibodies already developed, the third specific aim will be to determine whether mouse intestinal mucosal mast cells in vivo have the same phenotype as bone marrow culture-derived mast cells. Finally, the expression of one neutral GSL, globoside, appears to be associated with the secretroy granules of bone marrow culture- derived mast cells. The fourth specific aim is to determine whether or not this GSL is selectively located on perigranular membranes of these cells and to follow the movement of this molecule during mast cell activation. Because it is likely that the various mast cell populations and their knowledge of mast cell differentiation and heterogeneity is needed. It is anticipated that successful completion of the proposed research will yield useful new information about mast cell heterogeneity at both the intercellular and intracellular level.